A Sensitive Fluorescent Assay for Tetracycline Detection Based on Triple-helix Aptamer Probe and Cyclodextrin Supramolecular Inclusion

Abstract

Herein, an effective pyrene excimer signaled fluorescent biosensor for the determination of tetracycline based on triple-helix aptamer probe (TAP) and supramolecular inclusion of cyclodextrin was reported. The TAP was devised containing an aptamer loop, two DNA segment stems and a triplex-forming oligonucleotide (signal probe) labeled with pyrenes at 5′ and 3′ ends. The presence of target could result in its binding towards aptamer with a mighty affinity, leading to a conformation change of the TAP and whereupon the release of the signal probe. This liberty of signal probe enabled the formation of pyrene excimer, generating fluorescence signals. Further, signal amplification was fulfilled through the addition of γ-cyclodextrin which could interact with pyrene dimer, thus leading to an enhanced “on-state” of the sensing ensemble. In contrast, when the target was absent, the sensing ensemble remained “off-state” because of the long distance between two pyrene molecules. When the conditions were properly optimized, the increasing signal kept a linear dependence on target concentrations ranging from 5.0 nM to 100 nM, and the detection limit reached as low as 1.6 nM. In this way, a newly-constructed, simple, and economically affordable protocol enjoys desirable efficiency, sensitivity, specificity in biosensing. Also, its universality as another attractive behalf in assaying diverse targets was envisioned with only the need of matched aptamer replacement.